The polysaccharide starch constitutes one of the most important storage substances in plants. Starch is widely used for the production of foodstuffs and plays also a significant role as a regenerative raw material in manufacturing of industrial products. In order to use starches in many different technical areas a large variety of optionally modified starches is required in order to meet the varying needs of the processing industry.
Although starch consists of a chemically homogeneous basic component, namely glucose, it does not constitute a homogeneous raw material. It is a complex mixture of molecules which differ in their degree of polymerization and degree of branching of the glucose chains: Amylose-type starch is a basically unbranched polymer consisting of α-1,4-glycosidically branched glucose molecules, whereas amylopectin-type starch is a mixture of branched glucose chains, comprising additionally α-1,6-glycosidic interlinkings.
The molecular structure of starch mainly depends on its degree of branching, the amylose/amylopectin ratio, the average chain-length, chain lenght distribution, and degree of phosphorylation, further determining the functional properties of the starch and the aequous solutions thereof. Important functional properties of the starch, resp., the aequous solutions thereof are, e.g., solubility, tendency to retrogradation, capability of film formation, viscosity, pastification (binding and gluing) properties, and cold resistance. Additionally, the size of the starch granules may also determine the suitability of the starch for particular applications.
Since starch is often adapted by chemical and/or physical modification in order to meet the requirements of industry, there is a great need for the provision of modified starches which would render plant cells or plant parts containing modified starch more suitable for industrial processing, e.g., the production of foodstuff or technical products. Therefore, it is desired to avoid chemical and/or physical modification, which is time-consuming and expensive and to provide plants which synthesize a starch which meets more closely the demands of the starch processing industry.
Conventional methods for the preparation of modified plants which produce modified products, e.g., by classical breeding and/or the production of mutants, are limited to the use of homologous genes and are not always satisfying. Particularly in wheat, it is difficult to prepare a stable mutant by classical breeding due to the polyploidity of wheat (tetra- or hexaploidity). However, a wheat mutant producing waxy-type starch (amylose-free starch) was recently achieved by breeding methods (Nakamura et al., Mol. Gen. Genet. 248 (1995), 253–259).
A further alternative is the preparation of transgenic plants which comprise nucleic acid molecules suitable to modify plant starch metabolism in order to synthesize a modified starch. Such plants are produced by means of recombinant molecular biological techniques and the introduction of homologous and/or heterologous nucleic acid molecules (e.g., coding regions, regulatory elements, introns), which interfere in starch metabolism. However, the application of recombinant molecular biological techniques requires the availability of suitable nucleic acid which participate directly or indirectly (e.g., cosuppression, anti-sense-technology, generation of protein or ribozyme) in starch metabolism or starch biosynthesis (i.e., synthesis, modification and/or degradation of starch) with respect to quantity and/or quality of the starch.
Numerous genes are involved in starch metabolism. Therefore, a large number of genes encoding, e.g., branching enzymes, debranching enzymes, isoamylases, starch synthetases, ADP-glucose-pyrophosphorylases, have been used to modify starch metabolism in plants.
R1 proteins are involved in starch metabolism, especially with respect to the degree of phosphorylation of the starch and therefore, suitable to modify starch synthesis. In particular, R1-proteins and genes encoding R1-proteins derived from a number of plant species are known, i.e., potato from WO 97/11188-A1 and Lorberth et al., Nature Biotechnology 16 (1998), 473–477), maize from WO 98/27212-A1, rice from Sakaki et al., EMBL database entry Accession No. C 71741 (1997-09-19), and arabidopsis, ginger, mosses, cattail (Typha latifolia), and soybean from WO 99/53072-A1.
However, the presence of an R1-protein in wheat plants was not shown, corresponding nucleic acid molecules were not identified. Furthermore, the known nucleic acid molecules encoding R1-proteins are not always satisfying or suitable for the genetic engeneering or the in vivo mutagenesis of wheat plants in order to modify wheat starch biosynthesis and/or metabolism.